杀虫剂丁烯氟虫腈对家蚕卵巢培养细胞(BmN)的毒性试验  

Toxicity Test of Insecticide Butylene Fipronil on BmN Cells

作　　者：钟石[1] 朱俭勋[1] 李有贵[1] 王新全[2] 计东风[1] 

机构地区：[1]浙江省农业科学院蚕桑研究所,杭州310021 [2]浙江省农业科学院农产品质量标准研究所,杭州310021

出　　处：《蚕业科学》2016年第5期865-869,共5页ACTA SERICOLOGICA SINICA

基　　金：浙江省农药残留检测与控制研究重点实验室开放课题(No.2014ZPC03);农业部农药残留检测重点实验室开放课题(No.2014PRG03)

摘　　要：丁烯氟虫腈是在农业害虫防治中应用广泛的苯基吡唑类杀虫剂,以家蚕卵巢培养细胞（BmN）为材料,研究丁烯氟虫腈对家蚕的毒性及其作用机制。用MTT法检测丁烯氟虫腈对BmN细胞的增殖有较强抑制作用,且在药液质量浓度12.5~800μg/mL范围内,其抑制作用呈现出浓度依赖性,200、400μg/mL时抑制率分别高达41.34%±7.30%、56.40%±6.70%。用400μg/mL丁烯氟虫腈药液处理BmN细胞48 h后,用流式细胞仪检测发现可导致DNA合成期（S期）细胞比率显著提高（由33.51%±5.12%增加至44.33%±3.98%）,DNA合成前期（G0/G1）、DNA合成后期（G2/M）细胞比率降低（分别由37.31%±5.71%、29.18%±7.65%降至31.65%±2.33%、24.02%±3.66%）,细胞早期凋亡率和晚期凋亡率分别由4.74%±1.29%、4.95%±1.46%增加至9.86%±2.51%、25.37%±3.38%;用qRT-PCR法检测细胞中4种半胱氨酸蛋白酶（Caspase-1、Caspase-3、Caspase-4、Caspase-5A）基因mRNA转录水平显著上调。研究结果提示,丁烯氟虫腈可能通过上调BmN细胞的半胱氨酸蛋白酶基因表达水平,诱导BmN细胞S期阻滞和凋亡,从而抑制BmN细胞增殖。Butylene fipronil is a kind of widely used phenylpyrazole insecticide in control of agricultural pests. To study the toxicity and its functioning mechanism of butylene fipronil on silkworm（ Bombyx mori）,BmN cells were used as the material and MTT colorimetry was used to detect proliferation of BmN cells. It was found that BmN cells treated with bytulene fipronil exhibited significantly decreased cell proliferation in a dose-dependent manner within the range of 12. 5 to 800μg / mL. The inhibitory rate was 41. 34% ± 7. 30% and 56. 40% ± 6. 70% at the concentration of 200 and 400 μg / mL respectively. Flow cytometry was used to analyze cell cycle distribution and apoptosis of the treated cells. An evident Sphase arrest was observed in BmN cells treated with bytulene fipronil（ 400 μg / mL） for 48 h,with the population of Sphase cells increased from 33. 51 ± 5. 12% to 44. 33 ± 3. 98%,whereas the populations in the G0 / G1 and G2 / M phases markedly decreased from 37. 31 ± 5. 71% and 29. 18 ± 7. 65% to 31. 65 ± 2. 33% and 24. 02 ± 3. 66% respectively,and the rate of early and late apoptosis increased from 4. 74 ± 1. 29% and 4. 95 ± 1. 46% to 9. 86 ± 2. 51% and 25. 37 ± 3. 38%respectively. Moreover, expression of Caspase-1,Caspase-3, Caspase-4 and Caspase-5A genes was significantly increased in the bytulene fipronil-treated BmN cells as revealed by qRT-PCR. These results suggest that bytulene fipronil induces S-phase cell cycle arrest and apoptosis of BmN cells through up-regulation of caspase gene expression and thus inhibits proliferation of BmN cells.

关 键 词：杀虫剂 丁烯氟虫腈 BmN细胞 增殖 细胞周期 凋亡 半胱氨酸蛋白酶 

分 类 号：S884.96[农业科学—特种经济动物饲养] S482.3[农业科学—畜牧兽医]